Patients diagnosed with Anaplastic Large Cell Lymphoma (ALCL) are still treated with toxic multi-agent chemotherapy and as many as 25-50% of patients relapse. To understand disease pathology and to uncover novel targets for therapy, Whole-Exome Sequencing (WES) of Anaplastic Lymphoma Kinase (ALK)+ ALCL was performed as well as Gene-Set Enrichment Analysis. This revealed that the T-cell receptor (TCR) and Notch pathways were the most enriched in mutations. In particular, variant T349P of NOTCH1, which confers a growth advantage to cells in which it is expressed, was detected in 12% of ALK+ and ALK- ALCL patient samples. Furthermore, we demonstrate that NPM-ALK promotes NOTCH1 expression through binding of STAT3 upstream of NOTCH1. Moreover, inhibition of NOTCH1 with γ-secretase inhibitors (GSIs) or silencing by shRNA leads to apoptosis; co-treatment in vitro with the ALK inhibitor Crizotinib led to additive/synergistic anti-tumour activity suggesting this may be an appropriate combination therapy for future use in the circumvention of ALK inhibitor resistance. Indeed, Crizotinib-resistant and sensitive ALCL were equally sensitive to GSIs. In conclusion, we show a variant in the extracellular domain of NOTCH1 that provides a growth advantage to cells and confirm the suitability of the Notch pathway as a second-line druggable target in ALK+ ALCL.

A 1 Virtanen Institute for Molecular Sciences University of Eastern Finland Finland

Belarusian Centre for Paediatric Oncology Hematology and Immunology Minsk Belarus

Center of Molecular Medicine CEITEC Masaryk University Brno Czech Republic

College of Applied Medical Sciences King Abdulaziz University Jeddah Saudi Arabia

Colleges of Medicine and Applied Medical Sciences University of Hail Hail Saudi Arabia

Colleges of Medicine and Applied Medical Sciences University of Haill Haill Saudi Arabia

Department of Biochemistry University of Cambridge Tennis Court Road Cambridge UK

Department of Internal Medicine Hematology and Oncology University Hospital Brno Czech Republic

Department of Paediatric Oncology Addenbrooke Hospital Cambridge UK

Department of Pathology and Laboratory Medicine Cornell University New York NY USA

Department of Pathology Hematopathology Section UKSH Campus Kiel Kiel Germany

Department of Pathology Medical University of Vienna Vienna Austria

Department of Pathology University of Cambridge Cambridge UK

Department of Pediatric Hematology Oncology Hannover Medical School Hannover Germany

Diagnostic and Research Institute of Pathology Medical University of Graz Graz Austria

Institut Universitaire du Cancer Toulouse Oncopole et Université Paul Sabatier Toulouse France

Medical University of Vienna and Ludwig Boltzmann Institute for Cancer Research Vienna Austria

Our Lady Children Hospital Crumlin Ireland

Perelman School of Medicine Philadelphia USA

Unit of Laboratory Animal Pathology University of Veterinary Medicine Vienna Vienna Austria

University Hospital Hamburg Eppendorf Pediatric Hematology and Oncology Hamburg Germany

University of Milano Bicocca Monza Italy

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